Journal: Molecules

Article Title: Application of Cinnamomum burmannii Essential Oil in Promoting Wound Healing

doi: 10.3390/molecules29092080

Figure Lengend Snippet: BEO attenuated the inflammatory response in RAW 264.7 cells stimulated by LPS. ( A ) qPCR of IL-1β, IL-6 and TNF-α levels in RAW 264.7 cells that were stimulated by LPS at 24 h from different groups. ( B ) The level of IL-6 and TNF-α secreted by RAW 264.7 cells treated with BEO was determined by ELISA. ( C ) Effects of 150 μg/mL, 200 μg/mL, 250 μg/mL and 300 μg/mL BEO on the expression of NF-κB/p-IκBα protein. ( D ) The cell typing of RAW 264.7 was detected by flow cytometry. Polarization rate ( D ) and sectional images ( E , F ) of each group are shown. n = 3. *, p < 0.05; **, p < 0.01; ***, p < 0.001; **** p < 0.0001; ns, not significant.

Article Snippet: Monoclonal antibodies against β-actin, and NF-κB p65 were purchased from MedChemExpress Inc. (MCE, Monmouth Junction, NJ, USA).

Techniques: Enzyme-linked Immunosorbent Assay, Expressing, Flow Cytometry

Journal: Frontiers in Nutrition

Article Title: Cannabis oil modulates liver alterations and endocannabinoid system changes in a female rat model of diet-induced MASLD

doi: 10.3389/fnut.2026.1770150

Figure Lengend Snippet: Liver NrF2 and NFκB p65 expression in female rats fed a reference diet (RD), sucrose-rich diet (SRD) or SRD with cannabis oil (SRD + Ca). (A) Representative photomicrographs of immunocytochemical staining NrF2 in the liver sections of rats. Decreased levels of nuclear and cytoplasmic positive markers are observed in the SRD group. Scale bar 50 μm. (B) Quantitative immunohistochemical analysis of liver NrF2 expression expressed as integrated optical density (IOD). (C) Representative photomicrographs of immunocytochemical staining NFκB p65 in the liver sections of rats. Increased levels of nuclear and cytoplasmic positive markers are observed in the SRD group. Scale bar 50 μm. (D) Quantitative immunohistochemical analysis of NFκB p65 expression in the liver. Data are expressed as mean ± SEM ( n = 6). Statistical differences were evaluated by one-way ANOVA followed by the Newman–Keuls post hoc test (* P < 0.05).

Article Snippet: The samples were incubated in a humid chamber first with a specific primary antibody for 4-HNE (mouse monoclonal antibody; Catalog # MAB3249; R&D Systems), NrF2 (mouse monoclonal antibody; sc-365949; Santa Cruz Biotechnology), NF-κB p65 (mouse monoclonal antibody; sc-8008; Santa Cruz Biotechnology) and TGF-β1 (mouse monoclonal antibody; sc-52893; Santa Cruz Biotechnology) (for 14–16 h at 4°C) and then with biotin-conjugated secondary antibody (anti-mouse, 1:100 dilution, Sigma) for 30 min at room temperature.

Techniques: Expressing, Cannabis, Staining, Immunohistochemical staining

Journal: Journal of cell science

Article Title: The laminin-binding integrins regulate nuclear factor κB-dependent epithelial cell polarity and inflammation.

doi: 10.1242/jcs.259161

Figure Lengend Snippet: Fig. 7. α3−/−α6−/−collecting duct cells exhibit increased NF-κB activation. (A) α3f/fα6f/f and α3−/−α6−/−collecting duct cells were grown to 80% confluence. Then, 24 h later, conditioned medium from these cells was collected and subjected to cytokine multiplex analysis. A subset of cytokines with significant differences between α3−/−α6−/−and α3f/fα6f/f collecting duct cells (P<0.01) are shown. (B-D) α3f/fα6f/f and α3−/−α6−/−collecting duct cells grown to 80% confluence were lysed and subjected to separation of nuclear and cytosolic fractions. Western blot analysis of the fractions (20 µg total protein/lane) showed increased expression of nuclear p65 and cytosolic phospho-p65S536 (p-p65S536) in α3−/−α6−/−compared to α3f/fα6f/f collecting duct cells. Lamin A/C was used as a nuclear marker; β-actin was used as a loading control (B). The expression of p65 (C) or p-p65S536 (D) was normalized to β-actin and quantified as a ratio of α3f/fα6f/f cells. (E) α3f/fα6f/f and α3−/−α6−/−collecting duct cells were grown to 80% confluence. Then, 24 h later, NF-kB activity in these cells was detected using a Nano-Glo Luciferase Assay and quantified as a ratio of α3f/fα6f/f collecting duct cells. (F) α3f/fα6f/f and α3−/−α6−/−collecting duct cells grown to 80% confluence were treated with solvent (3% Tween 80 in regular growth medium) or 10 µM BMS-345541 in solvent for 1 h. NF-kB activity was detected using a Nano-Glo Luciferase Assay and quantified as a ratio of α3f/fα6f/f collecting cells. (G,H) Immunostaining for the collecting duct marker DBA (green) and phospho-p65S276

Article Snippet: In some experiments, the collecting duct cells were treated with IκB kinase inhibitor BMS-34554 (100 μM in water/3% Tween 80) or the vehicle alone for 1 h. To silence NFκB p65, collecting duct cells were transfected with non-silencing siRNA (20 nM, transfection control) or p65-silencing siRNA [NFκB p65 siRNA (m), 29411, Santa Cruz Biotechnology] using Lipofectamine RNAiMAX according to manufacturer’s instructions.

Techniques: Activation Assay, Multiplex Assay, Western Blot, Expressing, Marker, Control, Activity Assay, Luciferase, Solvent, Immunostaining

Journal: Journal of cell science

Article Title: The laminin-binding integrins regulate nuclear factor κB-dependent epithelial cell polarity and inflammation.

doi: 10.1242/jcs.259161

Figure Lengend Snippet: Fig. 8. The mesenchymal phenotype of α3−/−α6−/−collecting duct cells is mediated by NF-κB-dependent Snail and Slug induction. (A-C) α3f/fα6f/f and α3−/−α6−/−collecting duct cells grown to 80% confluence were lysed and subjected to separation of nuclear and cytosolic fractions. Western blot analysis of the fractions (20 µg total protein/lane) showed increased nuclear Snail and Slug in α3−/−α6−/−collecting duct cells compared to α3f/f/α6f/f collecting duct cells. Lamin A/C was used as a nuclear marker; β-actin was used as a loading control (A). The expression of Snail (B) or Slug (C) was normalized to β-actin and quantified as a ratio of α3f/fα6f/f cells. **P<0.01 between α3−/−α6−/−and α3f/fα6f/f collecting duct cells. (D-H) α3−/−α6−/−collecting duct cells grown to 80% confluence were transfected with non-silencing (n) or NF-κB p65-silencing (p65) siRNA for 48 h. Collecting duct cells were lysed and subjected to separation of nuclear and cytosolic fractions. Western blot analysis of the fractions (20 µg total protein/lane) showed decreased cytosolic and nuclear phospho-p65S536 (p-p65), total p65 and Slug, and decreased nuclear Snail in cells transfected with p65 siRNA compared to non-silencing siRNA. Lamin A/C was used as a nuclear marker; β-actin was used as a loading control (D). The expression of p-p65 (E), p65 (F), Snail (G) or Slug (H) was normalized to β-actin and quantified as a ratio of cells transfected with non-silencing siRNA. **P<0.01 between cells transfected with non-silencing and p65 siRNA. (I-M) α3f/f/α6f/f and α3−/−α6−/−collecting duct cells grown to 80% confluence were transfected with non-silencing or NF-κB p65-silencing (p65) siRNA, and lysed 48 h later. Total cell lysates (40 µg total protein/lane) were analyzed by western blotting for levels of p65, E-cadherin, N-cadherin and collagen I; β-actin was used as a loading control (I). The expression of p65 (J), E-cadherin (K), N-cadherin (L) or collagen I (M) proteins was normalized to actin and quantified as ratio of α3f/fα6f/f cells transfected with non-silencing siRNA. Data are mean±s.e.m. of at least three experiments. **P<0.01 between non-silencing and p65 siRNA-transfected α3f/f/α6f/f or α3−/−/α6−/−collecting duct cells. Statistical significance was determined using two-tailed, unpaired t-tests.

Article Snippet: In some experiments, the collecting duct cells were treated with IκB kinase inhibitor BMS-34554 (100 μM in water/3% Tween 80) or the vehicle alone for 1 h. To silence NFκB p65, collecting duct cells were transfected with non-silencing siRNA (20 nM, transfection control) or p65-silencing siRNA [NFκB p65 siRNA (m), 29411, Santa Cruz Biotechnology] using Lipofectamine RNAiMAX according to manufacturer’s instructions.

Techniques: Western Blot, Marker, Control, Expressing, Transfection, Two Tailed Test

Journal: Frontiers in Nutrition

Article Title: Astaxanthin alleviates altered hepatic lipid metabolism and oxidative stress in animals fed a high-sucrose diet

doi: 10.3389/fnut.2026.1781406

Figure Lengend Snippet: (A) Representative photomicrographs of p-NFκB p65 immunocytochemical staining in liver tissues (400 × magnification), and (B) quantitative immunohistochemical analysis of p-NFκB p65 expression in the liver of rats fed a reference diet (RD), RD plus ASTX (RD + ASTX), high-sucrose diet (HSD), and HSD plus ASTX (HSD + ASTX). Values (mean ± SEM, n = 6 ) are expressed as the in tegrated optical density (IOD). Values that do not share the same letter were significantly different ( p < 0.05) when one variable at a time was compared by two-way ANOVA. D, effect of Diet. I, effect of intervention. Int (D x I), interaction between diet x intervention. ns, not significant.

Article Snippet: The samples were incubated in a humid chamber first with a specific primary antibody for Nrf2 (mouse monoclonal antibody; sc-365949; Santa Cruz Biotechnology) and p-NFκB p65 (mouse monoclonal antibody; sc-136548; Santa Cruz Biotechnology) (for 14–16 h at 4 °C) and then with biotin-conjugated secondary antibody (antimouse, 1:100 dilution, Sigma) for 30 min at room temperature.

Techniques: Staining, Immunohistochemical staining, Expressing

Journal: Frontiers in Nutrition

Article Title: Astaxanthin alleviates altered hepatic lipid metabolism and oxidative stress in animals fed a high-sucrose diet

doi: 10.3389/fnut.2026.1781406

Figure Lengend Snippet: Mechanisms by which ASTX modulates liver lipid metabolism and oxidative stress in HSD-fed rats. ASTX supplementation mitigates hepatic injury by reducing steatosis and triglyceride accumulation, mainly through enhanced mitochondrial fatty acid β-oxidation (↑ CPT-1 activity) without affecting lipogenesis. In parallel, ASTX improves hepatic redox homeostasis by lowering ROS levels, increasing antioxidant enzyme activities (CAT, GST) and restoring GSH content. Increased Nrf2 and decreased p-NFκB p65 protein expression were also modulated by ASTX treatment. Solid lines indicate effects supported by the experimental data, whereas dashed lines represent suggested pathways not directly evaluated in this study.

Article Snippet: The samples were incubated in a humid chamber first with a specific primary antibody for Nrf2 (mouse monoclonal antibody; sc-365949; Santa Cruz Biotechnology) and p-NFκB p65 (mouse monoclonal antibody; sc-136548; Santa Cruz Biotechnology) (for 14–16 h at 4 °C) and then with biotin-conjugated secondary antibody (antimouse, 1:100 dilution, Sigma) for 30 min at room temperature.

Techniques: Activity Assay, Expressing

Journal: Cancer Science

Article Title: Targeting nuclear factor‐κ B suppresses the negative effect of T oll‐like receptor 4 signaling on antimetastasis therapy based on targeting αvβ3

doi: 10.1111/j.1349-7006.2012.02299.x

Figure Lengend Snippet: Toll‐like receptor 4 (TLR4) ligand promotes invasive migration of B16 cells. (a) Promoting effect of TLR4 ligands on invasive migration of B16 cells. B16 cells were untreated or treated with LPS (100 ng/mL) or molecules from damaged tumor cells (DTC‐Ms) (5, 20, 100 μg/mL) for 48 h. The cells were then used in Matrigel invasion assay. (b) Downregulation of TLR4 in B16 cells. Expression of TLR4 in untransfected cells (control) and transfected cells (control shRNA or TLR4 shRNA) was detected by Western blot. (c,d) B16 cells were used in Matrigel invasion assay after the indicated treatment. (c) Control B16 cells and TLR4‐knockdown B16 cells were cultured for 48 h in the absence or presence of LPS or DTC‐Ms (100 μg/mL). (d) B16 cells were cultured for 48 h in the absence or presence of LPS, 6‐amino‐4‐(4‐phenoxyphenylethylamino) quinazoline (QNZ; 20 nM), resveratrol (Res; 30 μM), SB203580 (SB; 20 μM), or wortmannin (WT; 10 nM). (e) Activity of nuclear factor‐κB (NF‐κB) in B16 cells. B16 cells were transfected with NF‐κB–luciferase reporter plasmid, and treated with LPS or DTC‐Ms in the absence or presence of QNZ. Activity of NF‐κB was detected by measuring the relative activity of luciferase (defined by relative light units). (f) Downregulation of NF‐κB p65 in B16 cells. Expression of NF‐κB p65 in untransfected cells (control) and transfected cells (control shRNA or NF‐κB p65 shRNA) was detected by Western blot. (g,h) B16 cells were used in Matrigel invasion assay after the indicated treatment. (g) Control B16 cells and p65‐knockdown B16 cells were cultured for 48 h in the absence or presence of LPS or DTC‐Ms. (h) B16 cells were cultured for 48 h in the absence or presence of recombinant CBD‐HepII polypeptide of fibronectin (CH50; 20 μg/mL), LPS, DTC‐Ms, and QNZ. *P < 0.05; **P < 0.01.

Article Snippet: For downregulation of TLR4 or NF‐κB p65 subunit, B16 cells were transduced with TLR4 shRNA or NF‐κB p65 shRNA lentiviral particles, or control shRNA lentiviral particles (Santa Cruz Biotechnology, Santa Cruz, CA, USA) according to the manufacturer's protocol.

Techniques: Migration, Invasion Assay, Expressing, Control, Transfection, shRNA, Western Blot, Knockdown, Cell Culture, Activity Assay, Luciferase, Plasmid Preparation, Recombinant

Journal: Cellular signalling

Article Title: Minocycline inhibits PDGF-BB-induced human aortic smooth muscle cell proliferation and migration by reversing miR-221- and -222-mediated RECK suppression.

doi: 10.1016/j.cellsig.2019.01.014

Figure Lengend Snippet: Fig. 4. Minocycline inhibits PDGF-BB-induced NF-κB and AP-1 activation. A, PDGF-BB induces time-dependent NF-κB activation. Quiescent SMC were incubated with PDGF-BB (10 ng/ml). At the indicated time periods, activation of NF-κB was analyzed by immunoblotting using antibodies that specifically detect phos- phorylated p65 at Ser536. B, PDGF-BB induces time-dependent AP-1 activation. Quiescent SMC incubated as in A were analyzed for AP-1 activation by im- munoblotting using antibodies that specifically detect phosphorylated c-Jun at Ser73. C, Silencing IKKβ or pre-treatment with minocycline inhibit PDGF-BB-induced NF-κB activation. SMC incubated with lentiviral IKKβ shRNA (moi0.5 for 48 h) were made quiescent and treated with PDGF-BB (10 ng/ml for 30 min). In a subset of experiments, quiescent SMC were incubated with minocycline (10 μM for 15 min) and then treated with PDGF-BB (10 ng/ml for 30 min). Activation of NF-κB was analyzed as in A. D, Silencing JNK2 or pre-treatment with minocycline inhibits PDGF-BB-induced AP-1 activation. SMC incubated with lentiviral JNK2 shRNA (moi 0.5 for 48 h) were made quiescent and treated with PDGF-BB (10 ng/ml for 30 min). In a subset of experiments, quiescent SMC were incubated with minocycline (10 mM for 15 min) and then treated with PDGF-BB (10 ng/ml for 30 min). Activation of AP-1 was analyzed as in B. Silencing IKKβ and JNK2 was confirmed by immunoblotting. JNK2 and IKKβ served as off-targets in IKKβ and JNK2 silenced cells, respectively (right hand panels in C and D). Tubulin served as a loading control. Bar graphs at the bottom of panels in A-D represent densitometric analyses from three independent experiments. ⁎P < .05 control, †P < .05 versus PDGF- BB (n = 3).

Article Snippet: The following lentiviral shRNA vectors against IKKβ (#sc-35645-V), p65 (#sc-29410-V), JNK2 (#sc-39101-V), and c-Jun (#sc-29223-V) were purchased from Santa Cruz Biotechnology, Inc. Lentiviral shRNA against RECK (SHCLNV-NM_021111; TRCN0000376461) and eGFP (SHC005V) were purchased from Sigma-Aldrich.

Techniques: Activation Assay, Incubation, Western Blot, shRNA, Control

Journal: Cellular signalling

Article Title: Minocycline inhibits PDGF-BB-induced human aortic smooth muscle cell proliferation and migration by reversing miR-221- and -222-mediated RECK suppression.

doi: 10.1016/j.cellsig.2019.01.014

Figure Lengend Snippet: Fig. 5. PDGF-BB induces SMC proliferation in part via miR-221 and miR-222. A, B, PDGF-BB induces miR-221 (A) and miR-222 (B) expression via IKKβ, NF-κB, JNK and AP-1. Quiescent SMC incubated with PDGF-BB (10 ng/ml) for 1 h were analyzed for miR- 221 (A) and miR-222 (B) expression by TaqMan® Advanced miRNA assays. The results were normal- ized to corresponding U6 expression. In a subset of experiments, SMC were incubated with lentiviral IKKβ, p65, JNK2 or c-Jun shRNA (moi0.5 for 48 h), made quiescent and then treated with PDGF-BB. C, D, miR-221 and miR-222 mediate PDGF-BB-induced SMC migration (C) and proliferation (D), without affecting cell viability (E). SMC were transduced with miR-221 or miR-222 inhibitor prior to the ad- dition of PDGF-BB (10 ng/ml). Cell migration was analyzed after 18 h using transwell migration assays (C). Cell proliferation was analyzed after 48 h by CyQUANT® Cell Proliferation Assay (D). Cleaved caspase-3 levels, as an indicator of reduced cell via- bility, was analyzed after 8 h by immunoblotting using antibodies that detect both total and cleaved caspase-3 levels (E). Hydrogen peroxide (H2O2, 100 μM) served as a positive control. ⁎P < at least 0.05 versus control, †P < .05 versus PDGF-BB (n = 6).

Article Snippet: The following lentiviral shRNA vectors against IKKβ (#sc-35645-V), p65 (#sc-29410-V), JNK2 (#sc-39101-V), and c-Jun (#sc-29223-V) were purchased from Santa Cruz Biotechnology, Inc. Lentiviral shRNA against RECK (SHCLNV-NM_021111; TRCN0000376461) and eGFP (SHC005V) were purchased from Sigma-Aldrich.

Techniques: Expressing, Incubation, shRNA, Migration, Transduction, CyQUANT Assay, Proliferation Assay, Western Blot, Positive Control, Control

Journal: Oncotarget

Article Title: HTLV-1 Tax upregulates early growth response protein 1 through nuclear factor-κB signaling

doi: 10.18632/oncotarget.17699

Figure Lengend Snippet: ChIP assay was performed to determine which EGR1 promotor segments bind to p65 in TaxP cells ( A ). PCR products were subjected to electrophoresis on a 1.2% agarose gel. Results are averaged from three independent ChIP assays ( B ). Error bars show standard deviations. TaxP or MT2 cells were transfected with pNFκB-luc and p65 shRNA, and luciferase activity was measured after 48 h ( C ). TaxP or MT2 cells were transfected with EGR1-luc (E1) and p65 shRNA, and luciferase activity was measured after 48 h ( D ). TaxP or MT2 cells were transfected with pNFκB-luc ( E ) or EGR1-luck ( F ) and treated with Bay 11-7082. After 48 h, luciferase activity was measured. Results are shown as relative expression normalized to the control. * p < 0.01.

Article Snippet: To construct a plasmid expressing full-length Tax with an RFP tag, the Tax CDS was subcloned from pCMV-Bam-Tax into the Bgl II and Hind III sites of the pmCherry-C1 vector. pCMV-M22 Tax and pCMV-M47 Tax plasmids were kindly provided by Edward Harhaj (The University of Miami). p65 shRNA (sc-29410-SH) was purchased from Santa Cruz.

Techniques: Electrophoresis, Agarose Gel Electrophoresis, Transfection, shRNA, Luciferase, Activity Assay, Expressing, Control

Journal: Oncotarget

Article Title: HTLV-1 Tax upregulates early growth response protein 1 through nuclear factor-κB signaling

doi: 10.18632/oncotarget.17699

Figure Lengend Snippet: MT2 or TaxP cells were transfected with p65 shRNA for 48 h, and ACTB, EGR1, and p65 were detected by western blot ( A ). MT2 or TaxP cells were treated with or without 5 mM BAY 11-7082 for 48 h, and ACTB, EGR1, and p65, Tax, p-IκB, and IκB were detected by western blot ( B ). Jurkat cells were transfected with pNF-κB-luc and plasmids expressing wild-type Tax, M22 Tax, or M47 Tax for 48 h, and luciferase activity was measured ( C ). Jurkat cells were transfected with the indicated EGR1 promoter-luc plasmids and plasmids expressing wild-type Tax, M22 Tax, or M47 Tax for 48h, and luciferase activity was measured ( D ). Jurkat cells were transfected with plasmids expressing wild-type Tax, M22 Tax, or M47 Tax for 48 h and EGR1, CREB, p65, Tax, and ACTB were detected by western blot E. Jurkat cells were transfected with EGR1-luc plasmids and plasmids expressing wild-type Tax, M22 Tax, or M47 Tax for 48 h, and luciferase activity was measured F. * p < 0.01.

Article Snippet: To construct a plasmid expressing full-length Tax with an RFP tag, the Tax CDS was subcloned from pCMV-Bam-Tax into the Bgl II and Hind III sites of the pmCherry-C1 vector. pCMV-M22 Tax and pCMV-M47 Tax plasmids were kindly provided by Edward Harhaj (The University of Miami). p65 shRNA (sc-29410-SH) was purchased from Santa Cruz.

Techniques: Transfection, shRNA, Western Blot, Expressing, Luciferase, Activity Assay

Journal: Oncotarget

Article Title: HTLV-1 Tax upregulates early growth response protein 1 through nuclear factor-κB signaling

doi: 10.18632/oncotarget.17699

Figure Lengend Snippet: TaxP ( A ) or MT2 cells ( B ) were transfected with pNF-κB-luc and different quantities of pcDNA3.0-EGR1 for 48 h, and luciferase activity was measured. TaxP ( C ) or MT2 ( D ) cells were transfected with different quantities of pcDNA3.0-EGR1 for 48 h, and EGR1, p65, Tax, and ACTB were detected by western blot. TaxP cells were transfected with pcDNA3.0-EGR1 for 24 h, and p65, Lamin B, and EGR1 in the nuclei were analyzed by western blot ( E ) Relative levels of nuclear p65 were analyzed from three individual experiments. TaxP or MT2 cells were transfected with EGR1 siRNA for 48 h and EGR1 was detected by western blot ( F ) MT2 or TaxP cells were transfected with EGR1 siRNA and pNF-κB-luc for 48 h, and luciferase activity was detected ( G ) Schematic showing the positive feedback loop between EGR1 and NF-κB activation in HTLV-1-infected cells ( H ) * p < 0.01.

Article Snippet: To construct a plasmid expressing full-length Tax with an RFP tag, the Tax CDS was subcloned from pCMV-Bam-Tax into the Bgl II and Hind III sites of the pmCherry-C1 vector. pCMV-M22 Tax and pCMV-M47 Tax plasmids were kindly provided by Edward Harhaj (The University of Miami). p65 shRNA (sc-29410-SH) was purchased from Santa Cruz.

Techniques: Transfection, Luciferase, Activity Assay, Western Blot, Activation Assay, Infection

Journal: Molecular Medicine Reports

Article Title: MicroRNA-451 relieves inflammation in cerebral ischemia-reperfusion via the Toll-like receptor 4/MyD88/NF-κB signaling pathway

doi: 10.3892/mmr.2019.10587

Figure Lengend Snippet: Downregulation of microRNA-451 affects TLR4, MyD88 and NF-κB/p65 protein expression in an in vitro model of cerebral ischemia-reperfusion. (A) Reverse transcription-quantitative PCR was used to analyze the expression of microRNA-451. (Green to red: −2 to 2.) (B) A heat map was constructed to show the expression of factors including TLR4, MyD88 and NF-κB/p65 following transfection with anti-451. (C) MicroRNA-451 binding sites in the 3′ untranslated region of TLR4. (D) A luciferase reporter assay was used to determine the level of transcription from the TLR promoter following transfection with anti-451. (E) A network signal path revealed that TLR4 may be an important signaling pathway regulated by microRNA-451. (F) TLR4, (G) MyD88 and (H) NF-κB/p65 mRNA expression levels were determined by reverse transcription-quantitative PCR following transfection with anti-451. ## P<0.01 vs. negative. Negative, negative control; anti-451, anti-microRNA-451; TLR4, Toll-like receptor 4; MyD88, myeloid differentiation primary response protein MyD88; NF-κB/p65, nuclear factor-κΒ.

Article Snippet: At 24 h after transfection with anti-miRNA-451 and si-p65 (sc-422642, Santa Cruz Biotechnology, Inc.), cells were treated with 100 ng/ml lipopolysaccharide (LPS) for 6 h at 37°C.

Techniques: Expressing, In Vitro, Reverse Transcription, Real-time Polymerase Chain Reaction, Construct, Transfection, Binding Assay, Luciferase, Reporter Assay, Negative Control

Journal: Molecular Medicine Reports

Article Title: MicroRNA-451 relieves inflammation in cerebral ischemia-reperfusion via the Toll-like receptor 4/MyD88/NF-κB signaling pathway

doi: 10.3892/mmr.2019.10587

Figure Lengend Snippet: Downregulation of microRNA-451 affects MyD88 and NF-κB/p65 protein expression through TLR4 in cerebral ischemia-reperfusion. (A) TLR4, (B) MyD88 and (C) NF-κB/p65 protein expression was determined in an in vitro model of cerebral ischemia-reperfusion. (D) Western blot analysis of TLR4, MyD88 and NF-κB/p65 protein expression. (E) TLR4 protein expression was determined by immunofluorescence. Magnification, ×200. ## P<0.01 vs. negative. Negative, negative control; anti-451, anti-microRNA-451; TLR4, Toll-like receptor 4; MyD88, myeloid differentiation primary response protein MyD88; NF-κB/p65, nuclear factor-κΒ.

Article Snippet: At 24 h after transfection with anti-miRNA-451 and si-p65 (sc-422642, Santa Cruz Biotechnology, Inc.), cells were treated with 100 ng/ml lipopolysaccharide (LPS) for 6 h at 37°C.

Techniques: Expressing, In Vitro, Western Blot, Immunofluorescence, Negative Control

Journal: Molecular Medicine Reports

Article Title: MicroRNA-451 relieves inflammation in cerebral ischemia-reperfusion via the Toll-like receptor 4/MyD88/NF-κB signaling pathway

doi: 10.3892/mmr.2019.10587

Figure Lengend Snippet: Overexpression of miR-451 affects MyD88 and NF-κB/p65 protein expression through TLR4 in cerebral ischemia-reperfusion. (A) Reverse transcription-quantitative PCR was used to analyze the expression miR-451. (B) TLR4, (C) MyD88 and (D) NF-κB/p65 protein expression was determined in an in vitro model of cerebral ischemia-reperfusion. (E) Western blot analysis of TLR4, MyD88 and NF-κB/p65 protein expression. ## P<0.01 vs. negative. Negative, negative control; miR-451, miR-451; TLR4, Toll-like receptor 4; MyD88, myeloid differentiation primary response protein MyD88; NF-κB/p65, nuclear factor-κΒ.

Article Snippet: At 24 h after transfection with anti-miRNA-451 and si-p65 (sc-422642, Santa Cruz Biotechnology, Inc.), cells were treated with 100 ng/ml lipopolysaccharide (LPS) for 6 h at 37°C.

Techniques: Over Expression, Expressing, Reverse Transcription, Real-time Polymerase Chain Reaction, In Vitro, Western Blot, Negative Control

Journal: Molecular Medicine Reports

Article Title: MicroRNA-451 relieves inflammation in cerebral ischemia-reperfusion via the Toll-like receptor 4/MyD88/NF-κB signaling pathway

doi: 10.3892/mmr.2019.10587

Figure Lengend Snippet: Regulation of miRNA-451 affects inflammation in cerebral ischemia-reperfusion. The expression levels of (A) TNF-α, (B) IL-1β, (C) IL-6 and (D) IL-18 were determined following the overexpression of miR-451. The expression levels of (E) TNF-α, (F) IL-1β, (G) IL-6 and (H) IL-18 were determined following the downregulation of miR-451. ## P<0.01 vs. negative. Negative, negative control; miR-451, microRNA-451; anti-451, anti-microRNA-451; TLR4, Toll-like receptor 4; MyD88, myeloid differentiation primary response protein MyD88; NF-κB/p65, nuclear factor-κΒ; TNF-α, tumor necrosis factor-α; IL, interleukin.

Article Snippet: At 24 h after transfection with anti-miRNA-451 and si-p65 (sc-422642, Santa Cruz Biotechnology, Inc.), cells were treated with 100 ng/ml lipopolysaccharide (LPS) for 6 h at 37°C.

Techniques: Expressing, Over Expression, Negative Control

Journal: Molecular Medicine Reports

Article Title: MicroRNA-451 relieves inflammation in cerebral ischemia-reperfusion via the Toll-like receptor 4/MyD88/NF-κB signaling pathway

doi: 10.3892/mmr.2019.10587

Figure Lengend Snippet: Inhibition of TLR4 reduces the effect of microRNA-451 on TLR4, MyD88, NF-κB/p65 protein expression in cerebral ischemia-reperfusion. The expression level of (A) TLR4, (B) MyD88 and (C) NF-κB/p65 were determined following transfection with anti-451 and exposure to the TLR4 inhibitor TAK-242. (D) Western blot analysis of TLR4, MyD88 and NF-κB/p65 protein expression. The expression levels of (E) TNF-α, (F) IL-1β, (G) IL-6 and (H) IL-18 were determined following transfection with anti-451 and exposure to TAK-242. ## P<0.01 vs. negative; **P<0.01 vs. anti-451. Negative, negative control; anti-451, anti-microRNA-451; TLR4, Toll-like receptor 4; MyD88, myeloid differentiation primary response protein MyD88; NF-κB/p65, nuclear factor-κΒ; TNF-α, tumor necrosis factor-α; IL, interleukin.

Article Snippet: At 24 h after transfection with anti-miRNA-451 and si-p65 (sc-422642, Santa Cruz Biotechnology, Inc.), cells were treated with 100 ng/ml lipopolysaccharide (LPS) for 6 h at 37°C.

Techniques: Inhibition, Expressing, Transfection, Western Blot, Negative Control

Journal: Molecular Medicine Reports

Article Title: MicroRNA-451 relieves inflammation in cerebral ischemia-reperfusion via the Toll-like receptor 4/MyD88/NF-κB signaling pathway

doi: 10.3892/mmr.2019.10587

Figure Lengend Snippet: Inhibition of MyD88 reduces the effect of microRNA-451 on MyD88/NF-κB/p65 protein expression in cerebral ischemia-reperfusion. The expression level of (A) MyD88 and (B) NF-κB/p65 were determined following transfection with anti-451 and exposure to the MyD88 inhibitor ST2825. (C) Western blot analysis of MyD88 and NF-κB/p65 protein expression. The expression levels of (D) TNF-α, (E) IL-1β, (F) IL-6 and (G) IL-18 were determined following transfection with anti-451 and exposure to ST2825. ## P<0.01 vs. negative; **P<0.01 vs. anti-451. Negative, negative control; anti-451, anti-microRNA-451; MyD88, myeloid differentiation primary response protein MyD88; NF-κB/p65, nuclear factor-κΒ; TNF-α, tumor necrosis factor-α; IL, interleukin.

Article Snippet: At 24 h after transfection with anti-miRNA-451 and si-p65 (sc-422642, Santa Cruz Biotechnology, Inc.), cells were treated with 100 ng/ml lipopolysaccharide (LPS) for 6 h at 37°C.

Techniques: Inhibition, Expressing, Transfection, Western Blot, Negative Control

Journal: Molecular Medicine Reports

Article Title: MicroRNA-451 relieves inflammation in cerebral ischemia-reperfusion via the Toll-like receptor 4/MyD88/NF-κB signaling pathway

doi: 10.3892/mmr.2019.10587

Figure Lengend Snippet: Knockdown of NF-κB/p65 reduces the effect of microRNA-451 on NF-κB/p65 protein expression of cerebral ischemia-reperfusion. The expression level of (A) NF-κB/p65 as determined following transfection with anti-451 and depletion of NF-κB/p65. (B) Western blot analysis of NF-κB/p65 protein expression. The expression levels of (C) TNF-α, (D) IL-1β, (E) IL-6 and (F) IL-18 were determined following transfection with anti-451 and the knockdown of NF-κB/p65. ## P<0.01 vs. negative; **P<0.01 vs. anti-451. Negative, negative control; anti-451, anti-microRNA-451; si-p65, small interfering RNA targeting NF-κB/p65; NF-κB/p65, nuclear factor-κΒ; TNF-α, tumor necrosis factor-α; IL, interleukin.

Article Snippet: At 24 h after transfection with anti-miRNA-451 and si-p65 (sc-422642, Santa Cruz Biotechnology, Inc.), cells were treated with 100 ng/ml lipopolysaccharide (LPS) for 6 h at 37°C.

Techniques: Knockdown, Expressing, Transfection, Western Blot, Negative Control, Small Interfering RNA